Browsing by Author "Keret, Rafael"
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- ItemDevelopment of a protocol for the rapid in vitro establishment of Eucalyptus clones(Stellenbosch : Stellenbosch University, 2020-03) Keret, Rafael; Hills, Paul N.; Nakhooda, Muhammad; Stellenbosch University. Faculty of AgriSciences. Dept. of Plant Pathology.ENGLISH ABSTRACT: The practice of cultivating Eucalyptus species using micropropagation has found favour in the development of successful forestry plantation programs due to its effectiveness in generating large numbers of juvenile plant stocks. Natural phytohormones and their synthetic analogues, called plant growth regulators (PGRs), are typically included in growth media during micropropagation. These compounds may be used to stimulate certain physiological processes in plants, allowing, for example, the in vitro manipulation of plant cell growth and differentiation, depending on the concentrations and combinations of specific PGRs in the medium. The development of micropropagation programs for Eucalyptus spp. are often clone-specific, necessitating optimisation for each clone. There is thus an inherent need for the development of a standardized protocol permitting the in vitro establishment and proliferation of multiple Eucalyptus clones. In the present study, the sensitivity and responsiveness of three Eucalyptus grandis × Eucalyptus nitens clones to various PGRs were investigated for the different in vitro growth stages. They include auxins such as indole-3-acetic acid (IAA), indole-3-butyric acid (IBA) and 1-Naphthaleneacetic acid (NAA). Similarly, the cytokinins kinetin, 6-benzylaminopurine (BAP) and meta-topolin were tested. The effects of strigolactones on micropropagation were examined using the synthetic strigolactone analogue, GR24. Parameters such as explant multiplication, shoot elongation and rooting were examined for all clones. The explants were incubated on a reduced PGR maintenance medium, from which source material was derived for subsequent experimentation, enabling the growth stages to be investigated independently. The highest level of bud initiation and shoot growth was observed for all Eucalyptus clones when the growth medium was supplemented with 0.5 mg ℓ-1 of meta-topolin in combination with 0.1 mg ℓ-1 of IAA. Optimal elongation for all the clones was detected upon addition of meta-topolin at 0.05 mg ℓ-1 and IAA at 0.5 mg ℓ-1. The peak rooting response was obtained with 0.5 mg ℓ-1 of IBA for clones 2 and 3, whilst 0.029 mg ℓ-1 of GR24 in combination with 0.5 mg ℓ-1 of IAA elicited optimal rooting for clone 1. The most consistent rooting response across all clonal lines, however, was obtained with the GR24 treatment. Clone 2 was found to be the most responsive to in vitro stimulation by auxin and this observation was further probed via RT-qPCR. Expression of the genes encoding the auxin efflux and influx transporters PIN1 and AUX1, respectively, were analysed to assess whether clonal response could be linked to expression of a particular gene(s). Equal expression levels of PIN1:AUX1 were detected for clone 2. Clones 1 and 3, however, exhibited an expression profile whereby PIN1 transcript levels exceeded those of AUX1 when treated with 0.5 mg ℓ-1 of IAA. These expression profiles suggest that equivalent expression of PIN1:AUX1 correlated with greater responsiveness to exogenously supplied IAA for clone 2. The findings of this study thus suggest a novel approach for the rapid determination of in vitro responses of valuable Eucalyptus genotypes, by investigating culture growth stages and gene expression levels.
- ItemThe physiological and genetic underpinnings that influence the xylem properties of Eucalyptus grandis, when subjected to different levels of water availability(2024-03-01) Keret, Rafael; Hills, Paul Norman; Drew, David Michael